This application addresses broad Challenge Area (15) Translational Science and specific Challenge Topic, 15-AI-102: Develop diagnostics and drugs for multiple or extensively drug-resistant tuberculosis (MDR/XDR TB). Tuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), remains one of the world's deadliest infectious diseases. Co-infection with HIV plus the emergence of drug resistant strains have made TB both difficult and expensive to treat. In order to eradicate tuberculosis, new drugs are needed that will kill wild-type and drug-resistant strains of Mycobacterium tuberculosis. The major challenge in developing new TB drugs is to identify metabolic processes that are required for viability of the bacteria. The long-term goal of our work is to understand which processes are essential for survival of Mycobacterium tuberculosis. We will achieve this through the development of small molecule chemical tools which inhibit specific enzymes thought to be important and then examine their effect on bacterial survival. As we, and others, develop small molecule inhibitors of specific biochemical processes, we will begin to learn which pathways are essential for bacterial survival. These pathways can then become targets for therapeutic intervention. This proposal centers on developing inhibitors of the nonmevalonate pathway, the initial steps in isoprenoid biosynthesis. This pathway is essential in Mtb, but absent in humans. We focus on Dxr, or 1-deoxy-D-xylulose 5-phosphate reducto-isomerase, which is the first committed step in the nonmevalonate pathway. Our hypothesis is that small molecule inhibitors of Dxr will be effective in killing Mycobacterium tuberculosis. Dxr inhibitor development will be based on two approaches: (I) synthesis of lipophilic prodrugs and bioisosteres to enhance cell penetration and (II) structure-based design of Dxr inhibitors using the recent co-crystal structure of Mtb Dxr and fosmidomycin, a known Dxr inhibitor. The activity of compounds will be assessed through inhibition of Mtb Dxr activity, as well as efficacy against intact Mtb bacilli. From these studies, we will determine the role of Dxr, and thereby the nonmevalonate pathway, in mycobacterial survival and provide a platform for further lead molecule development. PUBLIC HEALTH RELEVANCE: Tuberculosis (TB) is one of the world's deadliest infectious diseases, and co-infection with HIV plus the emergence of drug-resistance have made TB both difficult and expensive to treat. In order to create new TB drugs, we need to understand how best to kill the organism that causes TB, Mycobacterium tuberculosis. This proposal outlines experiments that will help us understand if a particular pathway (the nonmevalonate pathway) can be used to kill this organism and help to cure TB.